Reorganization of Destabilized Nodes of Ranvier in βIV Spectrin Mutants Uncovers Critical Timelines for Nodal Restoration and Prevention of Motor Paresis

Abstract

Disorganization of nodes of Ranvier is associated with motor and sensory dysfunctions. Mechanisms that allow nodal recovery during pathological processes remain poorly understood. A highly enriched nodal cytoskeletal protein βIV spectrin anchors and stabilizes the nodal complex to actin cytoskeleton. Loss of murine βIV spectrin allows the initial nodal organization, but causes gradual nodal destabilization. Mutations in human βIV spectrin cause auditory neuropathy and impairment in motor coordination. Similar phenotypes are caused by nodal disruption due to demyelination. Here we report on the precise timelines of nodal disorganization and reorganization by following disassembly and reassembly of key nodal proteins in βIV spectrin mice of both sexes before and after βIV spectrin re-expression at specifically chosen developmental time points. We show that the timeline of nodal restoration has different outcomes in the PNS and CNS with respect to nodal reassembly and functional restoration. In the PNS, restoration of nodes occurs within 1 month regardless of the time of βIV spectrin re-expression. In contrast, the CNS nodal reorganization and functional restoration occurs within a critical time window; after that, nodal reorganization diminishes, leading to less efficient motor recovery. We demonstrate that timely restoration of nodes can improve both the functional properties and the ultrastructure of myelinated fibers affected by long-term nodal disorganization. Our studies, which indicate a critical timeline for nodal restoration together with overall motor performance and prolonged life span, further support the idea that nodal restoration is more beneficial if initiated before any axonal damage, which is critically relevant to demyelinating disorders. Nodes of Ranvier are integral to efficient and rapid signal transmission along myelinated fibers. Various demyelinating disorders are characterized by destabilization of the nodal molecular complex, accompanied by severe reduction in nerve conduction and the onset of motor and sensory dysfunctions. This study is the first to report reassembly of destabilized nodes with sequential improvement in overall motor performance. Our study reveals that nodal restoration is achievable before any axonal damage, and that long-term nodal destabilization causes irreversible axonal structural changes that prevent functional restoration. Our studies provide significant insights into timely restoration of nodal domains as a potential therapeutic approach in treatment of demyelinating disorders.

Myelin Repair: From Animal Models to Humans

“…During demyelination, the nodes of Ranvier are disorganized, altering nerve conduction. Following remyelination, the nodes are reorganized, allowing functional restoration, within a critical time window, consistent with the notion that node restoration is more beneficial if initiated before the occurrence of axonal damage (Saifetiarova et al, 2018). Nevertheless, the remyelination of damaged axons remains possible and may promote axonal recovery and, thus, neuronal survival (Schultz et al, 2017).…”

Impact of Auditory Experience on the Structural Plasticity of the AIS in the Mouse Brainstem Throughout the Lifespan

“…Free-floating slices were blocked in 4% goat serum and 0.3% (w/v) Triton X-100, 0.1% Tween 20 in PBS for a 1 h and then were incubated with primary antibody overnight at 4°C. The following primary antibodies were used: rabbit anti β4 spectrin (1:250) as previously described (Saifetiarova et al, 2018), mouse monoclonal anti-MAP2 (1:200; Millipore Cat# MAB3418, RRID:AB94856), mouse monoclonal anti-ankyrinG (AnkG; 1:100; UC Davis/NIH NeuroMab Facility Cat# 75-146, RRID:AB_10673030), mouse monoclonal anti-Kv1.2 (1:250; UC Davis/NIH NeuroMab Facility Cat# 75-008, RRID:AB_10673030), mouse monoclonal anti-Na-pan (1:100; Sigma-Aldrich Cat# S8809, RRID:AB_10673030), and rabbit polyclonal anti-c-Fos (1:100; Synaptic System Cat# 226003). After 3 washes with PBS containing 0.1% Tween 20, slices were incubated with different Alexa-488 goat anti-mouse IgG1 or 568 goat anti-mouse IgG2b or 568 goat anti-rabbit or 647 goat anti-guinea pig secondary antibodies (1:1000; Invitrogen) accordingly for 2 h at room temperature.…”

Severe Form of ßIV-Spectrin Deficiency With Mitochondrial Dysfunction and Cardiomyopathy—A Case Report

“…Prior to the exome analysis, a lysosomal storage disease had been suspected due to inclusions in the Schwann cells of myelinated axons seen in the electron microscopy ( Figure 1 ). Saifetiarova et al has detected similar inclusions and deformations of myelinated axons due to mutations in the SPTBN4 gene in mutant mice resulting in demyelination and neurodegeneration of central and peripheral axons ( 12 ). These findings correspond to the general demyelination seen in the MRI of index patient I. ßIV-spectrin seems to play a major role in the preservation of the integrity and myelination process of axons.…”

“…Mutations in the SPTBN4 gene, coding for ßIV-spectrin, lead to defective clustering of the AIS and the Nodes of Ranvier, disturbed axonal membrane polarity, as well as the inability to generate action potentials inducing neurodegeneration ( 5 , 12 , 13 ). These mutations have been identified in seven individuals by Knierim et al and Wang et al causing congenital myopathy, neuropathy and central deafness ( 14 , 15 ).…”